Malignant melanoma is the 6th most common cancer for men and women in the USA. Early detection and surgical resection of melanoma remains the mainstay of curative treatment of melanoma. However, detecting early melanoma and predicting prognosis is often challenging. Therefore, developing analytical methodologies to detect and identify biomarkers in easily accessible body fluid such as peripheral blood would be highly valuable for the early diagnosis and management of cancer patients. Blood-based biomarkers have several advantages over tissue-based biomarkers. Peripheral whole blood represents systemic processes including immune responses and cellular communications. Given the presence of tumor-inhibiting and tumor-promoting immune responses and inflammation in melanoma, and given the presence of circulating tumor cells, endothelial cells, and bone-marrow-derived cells and precursor cells in the blood, we hypothesize that blood-based biomarkers using peripheral blood cells from melanoma patients and high-throughput qRT-PCR gene expression methodology will provide new biomarkers for melanoma. Although genetic signatures of melanoma tissues and melanoma cells have been extensively studied, blood-based biomarker studies of melanoma that represent tumor-associated environment in the blood circulation have not been conducted. The goal of this proposal is to develop a protocol for blood biomarker profiling of melanoma using high- throughput gene expression analysis. In the preliminary experiments using microarray and qRT-PCR analyses, we have identified differentially expressed unique genes in the blood of melanoma patients, even from stage I/II melanoma patients. The genes we discovered are unique sets not previously described in cancer tissues or disease blood. In this proposal, we will apply biostatistical methodologies to generate signature models in Specific Aim 1, and will analyze fractions of blood to identify the cellular subsets responsible for the changes in candidate genes in Specific Aim 2. Our proposal will provide the first comprehensive study that analyzes blood-based genetic biomarkers of malignant melanoma. In the future, we will apply the signature models identified in this proposal into a more-extensive pool of melanoma patients to verify our data, into a population of other cancers (such as colon, lung cancers) and inflammatory diseases (such as rheumatoid arthritis) to validate and refine the specificity, and into a specific melanoma patients to further answer questions on specific biomarkers. These studies will be proposed in future R01 funding opportunities. Furthermore, the methodology is potentially useful for the diagnosis of a wide range of diseases, prediction of prognosis and treatment response, and risk assessment, thus the proposed studies in sample preparation and data analysis will be of wide interest. PROJECT NARRATIVE developing analytical methodologies to detect and identify biomarkers in easily accessible body fluid such as peripheral blood would be highly valuable for the early diagnosis and management of cancer patients. We hypothesize that blood-based biomarkers using peripheral blood cells from melanoma patients and high-throughput qRT-PCR gene expression methodology will provide new biomarkers for melanoma. We have identified unique sets of genes not previously described in cancer tissues or disease blood in the blood of malignant melanoma patients, and will apply biostatistical methodologies to the genes to generate signature models for malignant melanoma.